Mucosal modulation by LGG and R. gnavus specific tryptophan metabolites

LGG 和 R. gnavus 特异性色氨酸代谢物对粘膜的调节

基本信息

批准号：
9788244
负责人：
RONALDO PARAOAN FERRARIS
金额：
$ 38.75万
依托单位：
RUTGERS THE STATE UNIV OF NJ NEWARK
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-20 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9788244
关键词：
16S ribosomal RNA sequencing ARNT gene Ablation Acute Address Affect Affinity Agonist Animal Model Anti-inflammatory Aryl Hydrocarbon Receptor Bacteria Binding Biological Biological Assay Cell Compartmentation Cell Nucleus Cell physiology Colitis Collaborations Data Diet Disease Elements Engineering Epithelial Epithelial Cells Essential Amino Acids Excision Gene Expression Genes Genetic Germ-Free Gnotobiotic Health Homeostasis Immune Impairment Inflammatory disease of the intestine Innate Immune Response Intervention Intestinal Mucosa Intestines Knock-out Knockout Mice Lactobacillus Lactobacillus casei rhamnosus Lamina Propria Ligands Link Literature Mammals Mediating Mucolytics Mucosal Immune Responses Mucous Membrane Mus Nature Organoids Pathway interactions Perfusion Physiology Play Predisposition Probiotics Property Protein Biosynthesis Public Health Receptor Activation Receptor Signaling Recording of previous events Role Signal Pathway Signal Transduction Source Structure System Testing Tight Junctions Tryptamines Tryptophan Work aryl hydrocarbon receptor ligand bacterial genetics beta catenin cytokine dietary manipulation dimer dysbiosis experience high risk host-microbe interactions innovation metabolomics microbial microbial community microbiota mouse model mutant pathobiont promoter receptor structure function screening stem cell division transcriptome sequencing

项目摘要

PROJECT SUMMARY: The dietary benefits of tryptophan have been classically linked to its property as an essential amino acid required by mammals for protein synthesis, but recent work in the past decade has revealed its equally essential role as an important source of metabolites for the microbial community to generate signals, mainly via the Aryl Hydrocarbon Receptor (AHR), to affect a plethora of cellular functions. AHR signaling pathway is highly promiscuous not only responding to diverse ligands with resembling structures, but also crosstalk with and is modulated by numerous cellular signaling pathways. Distinct agonists can result in ligand selective differences in AHR structure and function, ultimately leading to agonist-selective gene expression and biological effects. In the gut, AHR activation by bacterial tryptophan catabolites influences mucosal homeostasis and microbiota composition. Although bacterial tryptophan metabolites and AHR axis may contribute to microbial sensing and tolerance, it is unclear if probiotics and pathobionts produce distinct tryptophan metabolites thereby eliciting bacterial species- and agonist-specific host mucosal modulating effects. Both Lactobacillus and Ruminoccocus spp catabolize tryptophan to a variety of tryptamine and indolic products that bind at low to high affinities to the AHR. The identities and specific biological effects of tryptophan metabolites produced by the probiotic Lactobacillus rhamnosus GG (LGG) and the pathobiont R. gnavus are currently unknown. Preliminary RNA-Seq analysis of mouse intestinal mucosa perfused with live LGG revealed a robust induction of AHR signaling activity as well as anti-inflammatory cytokines. Bacterial 16s rRNA sequencing and untargeted fecal metabolomics screening of a newly engineered Lyz1 knockout mouse model revealed a dysbiosis featured by expansion of mucolytic bacterial species (e.g., R. gnavus), accompanied by a pronounced elevation of tryptophan metabolites and a changed epithelial cell composition. The hypothesis is that the probiotic LGG and the pathobiont R. gnavus metabolize dietary tryptophan into distinct agonists that elicit differential mucosal modulating effects via the host AHR signaling. Aim I will use mono-colonization of germ-free mice, dietary tryptophan depletion, unbiased metabolomics profiling, and isogenic mutant bacteria deficient in tryptophan-metabolizing to identify LGG and R. gnavus specific tryptophan metabolites and their differential mucosal modulating effects. Aim 2 will employ organoid culture, intraluminal perfusion of AHR agonists, and genetic ablation of host AHR signaling to determine the direct effects of bacterial species-specific tryptophan metabolites and AHR signaling on Lyz1-deficient mouse mucosal homeostasis and colitis susceptibility. This MPI project uses rigorous approaches, including gnotobiotics, bacterial genetics, dietary manipulation, and unbiased metabolomics, etc. to address an important mechanism involving diet-microbe-host interaction that may underlie the biological natures of probiotics and pathobionts. The two PIs have a history of productive collaboration, and have demonstrated experiences in studying intestinal physiology. This project employing innovative animal models is significant for public health and disease intervention.

项目概要：色氨酸的饮食益处传统上与其作为必需氨基酸的特性有关哺乳动物用于蛋白质合成，但过去十年的最新研究揭示了其同样重要的作用微生物群落产生信号的重要代谢物来源，主要通过芳基碳氢化合物受体 (AHR)，影响多种细胞功能。 AHR 信号通路高度混杂不仅响应具有相似结构的不同配体，而且还与和发生串扰受多种细胞信号通路的调节。不同的激动剂可能导致配体选择性差异 AHR 结构和功能，最终导致激动剂选择性基因表达和生物效应。在肠道，细菌色氨酸分解代谢物激活 AHR 影响粘膜稳态和微生物群作品。尽管细菌色氨酸代谢物和 AHR 轴可能有助于微生物传感和耐受性，目前尚不清楚益生菌和致病菌是否产生不同的色氨酸代谢物，从而引发细菌种类和激动剂特异性宿主粘膜调节作用。乳酸菌和瘤胃球菌 spp 将色氨酸分解代谢为各种色胺和吲哚产物，这些产物以低到高的亲和力与阿赫。益生菌产生的色氨酸代谢物的特性和特定生物效应鼠李糖乳杆菌 GG (LGG) 和致病菌 R. gnavus 目前未知。初步RNA测序对灌注活 LGG 的小鼠肠粘膜的分析揭示了 AHR 信号活性的强烈诱导以及抗炎细胞因子。细菌 16s rRNA 测序和非靶向粪便代谢组学新设计的 Lyz1 敲除小鼠模型的筛选揭示了一种生态失调，其特征是粘液溶解细菌（例如 R. gnavus），伴有色氨酸代谢物明显升高以及上皮细胞组成的改变。假设益生菌 LGG 和致病生物 R. gnavus 将膳食色氨酸代谢为不同的激动剂，通过宿主引起不同的粘膜调节作用 AHR 信号。目标我将使用无菌小鼠的单一定植、膳食色氨酸消耗、无偏见代谢组学分析，以及色氨酸代谢缺陷的同基因突变细菌以鉴定 LGG 和 R。 gnavus 特异性色氨酸代谢物及其不同的粘膜调节作用。目标 2 将雇用类器官培养、AHR 激动剂的腔内灌注以及宿主 AHR 信号传导的基因消融以确定细菌物种特异性色氨酸代谢物和 AHR 信号对 Lyz1 缺陷小鼠的直接影响粘膜稳态和结肠炎易感性。该 MPI 项目采用严格的方法，包括悉生素、细菌遗传学、饮食控制和公正的代谢组学等，以解决一个重要的问题饮食-微生物-宿主相互作用的机制可能是益生菌生物学性质的基础致病生物。这两位 PI 有着富有成效的合作历史，并在以下领域展现了丰富的经验：研究肠道生理学。该项目采用创新动物模型对公共卫生具有重要意义和疾病干预。